Lecture Seamus Holden
|17 November 2010||FWN-Building 5116.0116, Nijenborgh 4, 9747 AG, Groningen|
|Speaker:||Dr. Seamus Holden|
|Affiliation:||University of Oxford, United Kingdom|
|Title:||Close to the edge: single-molecule FRET and imaging at the limits of resolution|
|Date:||Wed Nov 17, 2010|
|Host:||A. van Oijen|
|Telephone:||+31 50 363 9883|
The study of molecular machines such as the E. coli repair polymerase, DNA polymerase I (DNAP), is inherently a multiscale problem. On the nanoscale, we wish to understand DNAP as a single machine, characterised by distance fluctuations on the scale of one DNA base pair (0.34 nm). On the microscale we are interested in the spatial organisation of multiple DNAPs within the cell. This requires cutting-edge resolution in both regimes.
On the nanoscale, we extended the limits to resolution of smFRET carried out by total internal reflection fluorescence (TIRF) microscopy to the level of one DNA base pair, and characterised the major limiting factors to ultimate resolution.
On the microscale, we present a new approach (adapted from astronomy), which dramatically improves the performance of the algorithms used to analyse super-resolution microscopy data. Our approach improves accuracy, reduces image acquisition times and increases the ultimate spatial resolution of the technique. We are currently working towards application of these two methodological paradigms: to study DNAP translocation by smFRET, and intra-cellular DNAP spatial organisation by super-resolution imaging.
|Last modified:||22 October 2012 2.30 p.m.|